I. Field of the Invention
The present invention relates to the fields of biology and diagnostics. More specifically, the invention deals with “drop”-based technologies that permit simple, fast and accurate detection of most any analyte in environmental, plant, medical and other samples.
II. Related Art
A major stumbling block to low resource and/or home diagnostics is simplicity of design. Complex designs drive up the cost of manufacturing and limit distribution. But even clearing this hurdle is no guarantee, since complex designs also fail with improper use by those unskilled in their principles of operation. Thus, a diagnostic design that is inexpensive to manufacture, is simple to operate and can be readily interpreted by the unskilled end-user at home would be highly desirable. The desperate need for such an assay is well-demonstrated by the continuing health management issues surrounding malarial infections, with 250 million cases of malaria being reported in 2006, and having with a mortality rate of 881,000 individuals (FIG. 1) (WHO, 2006)
Traditionally, the “gold standard” for malarial detection has been microscopic examination of thick and/or thin blood films, where an experienced microscopist counts the number of parasites found per unit volume of blood (Warhurst & Williams, 1996). Other methods have also been developed, including fluorescence staining (Gay et al., 1996; Cooke et al., 1992; Srinivasan et al., 2000), amperometric immunosensing (Sharma et al., 2008), and single and multiplexed PCR techniques (Snounou et al., 1993). Recently, a new strategy using an enzyme-linked immunosorbent assay (ELISA) has been reported that detects the presence Histidine-Rich Protein 2 (pfHRP2), which is found specifically in P. falciparum induced malarial infection (Kifude et al., 2008; Martin et al., 2009). Although all of these techniques have limits of detection of <100 parasites/μl, they are limited primarily to the laboratory due to their sensitivity to environmental changes, requirements for specialized reagents, slow developing time, and reliance on sophisticated equipment for an interpretable readout. This is problematic since a majority of malarial infections occur in regions where advanced scientific technology and personnel are not readily available. Therefore, diagnostic strategies that are simple to use and require reagents and equipment that are easy to transport and stable in a variety of environmental conditions must be developed.
To circumvent these challenges, rapid diagnostic tests (RDTs) have been developed over the last two decades (Moody, 2002; Quintana et al., 1998; Singh et al., 2000). Although these techniques have performed well in laboratory testing, show high sensitivity, and acceptable limits of detection (<100 parasites/μL), these techniques have not performed well when subjected to variable temperature and humidity conditions due to their reliance on antibodies with small stability ranges (Ochola et al., 2006). In addition, there have been reports that mutations of pfHRP2 found in the Asia-Pacific region reduce the sensitivity of antigen detection due to the antibodies high specificity in the RDTs (Baker et al., 2005). Thus, even further improved and more effective diagnostic assays are needed.